Current available therapies for the management of severe pain, in its many forms, are largely inadequate. Acute pain, which typically persists for less than a month and is associated with a direct cause, such as tissue damage resulting from injury or surgery, can be distinguished from chronic pain, which is longer-lasting, occurring over months or years. Many chronic pain sufferers are especially poorly treated or are undertreated, which negatively impacts their quality of life and contributes to an inability to work, emotional distress, mood disorders, disrupted social integration and depression. Chronic pain often is characterized by short term episodic recurrences followed by an eventual relapse, only to later emerge again. Thus shorter-term treatments that are appropriate for acute pain therapy can be used with some success for chronic pain therapy when given in the midst of an episodic recurrence.
Opioids have been used to treat severe pain since antiquity. They remain the major class of pain relievers used today to treat post-surgical and cancer pain, largely due to of their effectiveness and rapid onset of pain relief. The opioid class of pain relievers interacts with specific receptors in the brain that are also found outside the CNS. The opioid family of receptors includes the mu (μ), kappa (κ) and delta (δ) receptors as well as the nociceptin receptor, also called the orphanin or ORL-1 receptor. Molecules that interact with these receptors have been classified by their ability to promote or block receptor signaling, and are commonly termed full agonists, partial agonists, mixed agonist-antagonists, or antagonists, depending upon the nature and consequence of their physical interactions with one or more of these receptors. The most effective opioid pain relievers are full agonists or partial agonists of either the μ or κ receptors, with μ receptor agonists (termed MOR agonists, for mu opioid receptor) most often used. For comprehensive reviews on the opioid receptors as analgesic targets see (1-3).
Opioid receptor agonists are most often used for treating severe and ongoing pain in humans and/or other mammals. Opioid analgesics that are effective in humans also produce antinociception in rodent laboratory models. Antinociception is defined as the blockade of the perception of a nociceptive stimulus, wherein a nociceptive stimulus is defined as one conferring “somatic or visceral pain processed by a normal, unaltered nervous system” (4). Two traditional tests of antinociception in mice and rats are the “hot plate” and “tail flick” assays, wherein the response latency following exposure to an uncomfortable thermal stimulus is monitored. Antinociception is evident when the test drug induces an extension of latency of response to the stimulus that proves to be significantly greater than the latency of the basal response. The hot plate test involves limited exposure of the paw pads to a heated surface and the animal is monitored for the time it takes to remove its paws from the surface. The tail flick test involves exposing the tail to a thermal stimulus and the latency for movement from the heat source of the tail is measured.
All marketed MOR agonists (e.g., morphine, oxycodone, meperidine, methadone, levorphanol, fentanyl), while effective in therapy for many types of pain, cause several unwanted side effects. These include constipation, respiratory depression, nausea, confusion, sedation, hypotension, development of tolerance (i.e., requiring higher and higher doses over time to maintain effective pain relief), dependence, and the prevalence of post-treatment withdrawal symptoms. Each of these effects contributes to an underutilization of these pain relievers for effective pain therapy. Often a minimum dose that is required to achieve adequate pain relief is near to the minimum dose that is required to elicit unwanted and possibly deadly side effects. This relationship, commonly termed a safety index or therapeutic window, differs significantly within the patient population. Its narrowness requires constant monitoring of patients receiving these drugs for signs of side effects, particularly those that are life-threatening (respiratory failure, as an example). Thus the duration of hospital stays is significantly increased for hospitalized patients treated with MOR agonists, with discharge often being delayed until doses of administered drug can be significantly lowered to the point at which normal bowel movements, signs of normal respiration and evidence of normal cognitive function become apparent.
The unwanted side effects of MOR agonists can be addressed to some extent by very careful dose titration and close patient monitoring. Morphine is often used as the opioid pain reliever of choice, for several reasons. It is relatively inexpensive and it has very well-characterized pharmacokinetics and pharmacodynamics that suggest doses for which the target therapeutic window is likely to be reached. Often, however, the necessary dose titration may take many hours or even days to establish, with inadequate pain relief obtained in the interim. Higher doses may be required following the onset of tolerance. The respiratory depression and constipation issues are perhaps the areas of highest concern, as the former can result in fatal overdose and the latter may be severe, debilitating, and require extended hospitalization or even require surgical intervention. Physical dependence is also a concern in long-term opioid therapy, as cessation of opioid drug administration may lead to apathy, weight loss, insomnia, anxiety, sexual dysfunction, and overwhelming drug cravings (1, 3). Dependence and addiction also contribute to societal concerns, such as compromised public safety owing to opiate-related crime.
The balance of the different μ-opioid-induced side effects differs slightly among known MOR agonists, but all such therapies have inherent risks. The severity of the side effects encountered are also generally proportional to the degree of pain relief provided by these drugs, which led to a simplistic model that viewed receptors as similar to an on/off switch, wherein an ‘on’ mode is characterized by receptor agonism (either full or partial) and an ‘off’ mode is characterized either by a lack of receptor agonism or by receptor antagonism. This model largely excluded the notion that the pain relieving properties of any opioid drug can theoretically be dissociated from one or more of its possible side effects. The model held that one switch, turned on, must result in a constellation of both positive and negative effects.
In recent years the physical structure of the MOR has been determined by X-ray techniques (5). Even prior to this achievement, the complexity of MOR signaling was becoming more apparent. There is not just one cascade of events, or signal, conveyed upon binding of a MOR agonist, but several. These downstream events include G protein coupling, adenylyl cyclase inhibition, receptor phosphorylation, βarrestin recruitment, and receptor internalization, events coordinated by regulatory proteins whose activity is dependent upon receptor activation (for reviews see (1, 6)). Small differences in the degree of activation of these different pathways may account for the slight differences in the side effect profiles among the different known MOR agonists. In addition, location of receptors, such as those on certain nerve terminal versus those in peripheral sites may also contribute to the side effect profiles. Drug potency and efficacy has usually been measured in vitro using assays for G protein coupling and for adenylyl cyclase inhibition. Assays measuring other downstream events, however, are not always in agreement with such potency assessments for certain MOR agonists.
Different G protein-coupled receptor agonists can in fact trigger distinct receptor signaling pathways. This phenomenon has been termed “functional selectivity”, “ligand-directed signaling”, “biased agonism” and “collateral efficacy” by the pharmacology community; such insights are strongly impacting new approaches in drug discovery (7-12). This concept is based on the idea that the chemical characteristics of the ligand may alter the conformation of the receptor such that it will interact preferentially with certain cellular proteins to mediate distinct biological responses. Therefore, the properties of the ligand bound dictate the nature of the receptor-protein interactions. Currently, marketed MOR agonists do not display significant functional selectivity. Moreover, there exists no a priori reason why such selectivity would not be possible. If different receptor signaling pathways ultimately lead to the manifestation of different side effects, a functionally selective MOR agonist can in principle be a potent pain relieving agent with an altered side effect profile, with one or more side effects being diminished or even nonexistent.
Over a decade ago the Bohn Lab began studying mice lacking GPCR regulatory proteins and found that mice with a genetic deletion of βarrestin2 display enhanced and prolonged morphine-induced antinociception (13-18). βArrestin2 is a scaffolding protein that can ad as a desensitizing element or as a signal transduction facilitator. Studies in the Bohn group have shown that morphine-induced analgesia is enhanced while tolerance is attenuated in mice lacking βarrestin2. Other studies in the Bohn group show that the severity of certain side effects, including physical dependence, constipation, and respiratory suppression are significantly reduced in mice lacking βarrestin2 (11, 14, 17-20). This suggests that in at least some organ systems and brain regions, βarrestin2 facilitates MOR signaling, that βarrestin2 plays a key role in determining MOR responsiveness, and that it serves as a critical switch in determining some of morphine's many effects. Therefore, agonists that could activate G protein-mediated signaling of the mu opioid receptor without inducing interactions with βarrestin2 would be a new class of potent pain relieving agents that should display fewer side effects. Such MOR agonists will promote antinociception, and based upon the animal model, should promote less respiratory suppression, dependence, tolerance and/or constipation. Since the publication of these findings and the realization of the possible therapeutic implications, attempts to discover such ligands have been a major interest within the pharmaceutical research community (21). The first such disclosed compound, TRV-130 (from Trevena, Inc.) is a “biased” MOR agonist and as of this writing is soon scheduled to enter Phase 2 clinical trials (22-24).
The desire to quantify functional selectivity has given rise to several mathematical models, all of which have been initially based on the Black and Leff operational model (Nobel Prize, 1988) (25). Development of this model has led to a means of simultaneously comparing the response that an agonist can elicit in one assay to the response elicited in another by normalizing its relative efficacy and potency to the performance a reference agonist that is run in parallel in both assays. Using the operational model allows one to simultaneously compare relative potencies and efficacies within an assay relative to the performance of a reference agonist, when properly normalized to minimize the contribution that differences in assay methodologies may impart. The relative efficiency of an agonist's ability to perform with respect to individual functions can then be compared. Reference compounds must be routinely used as comparators so that the potency and efficacy differences are not greatly influenced by inter-assay variation.
This usefulness of bias calculations is illustrated by the following example comparing relative potencies of an opioid enkephalin analog ([D-Ala2, N-MePhe4, Gly-ol]-enkephalin, “DAMGO”) and morphine in selected assay formats. In a βarrestin2 recruitment assay using a commercial enzyme fragment complementation protocol, DAMGO and morphine showed EC50 values of 217 and 361 nM, respectively. When an alternative βarrestin2-MOR assay was used, a bioluminescent resonance energy transfer (BRET) methodology, EC50 values of 19 nM (DAMGO) and 46 nM (morphine) nM were obtained (26). While the absolute values reported are markedly different in the two assay systems, the relative potency is in general agreement. A similar trend is seen in results of a G protein coupling assay, where DAMGO was shown to be roughly 2 fold more potent (EC50 values of 20 nM for DAMGO, 43 nM for morphine). In comparison to DAMGO, morphine is generally less efficient across assays; i.e., it is a partial agonist in both systems with a lower potency for stimulating both responses. Upon mathematical application of the operational model to both systems, normalized to the performance of DAMGO in both assays, one would calculate a “bias factor” of 1 for DAMGO (it is performing in par with itself) and a value approaching 1 for morphine (in our hands this value is 1.07). A truly biased ligand significantly diverges from 1, reflecting its ability to act on par with DAMGO in one assay, but less (or more) efficiently in another (27).